It is now well established that the transmission of pain information from the spinal cord to the brain can be inhibited through activation of a descending system. This system includes the area of periaqueductal gray (PAG), the medullary nuclei, nucleus raphe magnus (NRM) and nucleus magnocellularis (NMC). Physiological and behavioral experiments have shown that electrical stimulation of PAG or NRM and injection of morphine into the PAG can activate this system and produce strong analgesia. At the present time the mechanism by which this system is activated under natural conditions is not known. Recently, we have shown that injection of neurotensin (NT) into the PAG produces strong and prolonged analgesia. In this proposal we plan to determine: 1) if the analgesia produced by NT is due to its excitatory effect on the PAG neurons. Using single unit extracellular recording and micropressure injection techniques, we will examine the effect of NT on the PAG neurons that have direct projection to the NRM and/or NMC areas. 2) Using single unit recording techniques, we will determine if injection of NT into the PAG modifies the activity of those cells in the NRM and NMC that have direct projection to the spinal cord. 3) In order to explain the mechanism by which NT produces its long acting analgesic effect we plan to a) test the possibility that application of NT reduces the threshold of the cells in the PAG or NRM and/or NMC and causes these cells to be more readily excited by collateral of sensory afferents by using extracellular recording technique and b) by using intracellular recording techniques and bath application of NT in slice preparation of PAG, the duration of action of NT will be examined. 4) We will use single electrode voltage clamp techniques and noise analysis methods using an in vitro preparation to determine the kinetics of the membrane channels that are activated by NT. 5) Since it is possible that NT is the transmitter that under normal conditions leads to activation of the PAG cells, we plan to determine the location of neurotensinergic cells that have direct projection to the PAG. Double labeling with fluorescent dyes and immunocytochemical and autoradiographic methods will be used in this part of the experiment.